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MPT0B098, a Microtubule Inhibitor, Suppresses JAK2/STAT3 Signaling Pathway through Modulation of SOCS3 Stability in Oral Squamous Cell Carcinoma.

Peng HY, Cheng YC, Hsu YM, Wu GH, Kuo CC, Liou JP, Chang JY, Jin SL, Shiah SG - PLoS ONE (2016)

Bottom Line: In the present study, we examined the effect of a novel small-molecule microtubule inhibitor, MPT0B098, on STAT3 signaling in oral squamous cell carcinoma (OSCC).The accumulation of SOCS3 protein enhanced its binding to JAK2 and TYK2 which facilitated the ubiquitination and degradation of JAK2 and TYK2, resulting in a loss of STAT3 activity.Moreover, the combination of MPT0B098 with the clinical drug cisplatin or 5-FU significantly augmented growth inhibition and apoptosis in OSCC cells.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Cancer Research, National Health Research Institutes, Miaoli, Taiwan.

ABSTRACT
Microtubule inhibitors have been shown to inhibit Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signal transduction pathway in various cancer cells. However, little is known of the mechanism by which the microtubule inhibitors inhibit STAT3 activity. In the present study, we examined the effect of a novel small-molecule microtubule inhibitor, MPT0B098, on STAT3 signaling in oral squamous cell carcinoma (OSCC). Treatment of various OSCC cells with MPT0B098 induced growth inhibition, cell cycle arrest and apoptosis, as well as increased the protein level of SOCS3. The accumulation of SOCS3 protein enhanced its binding to JAK2 and TYK2 which facilitated the ubiquitination and degradation of JAK2 and TYK2, resulting in a loss of STAT3 activity. The inhibition of STAT3 activity led to sensitization of OSCC cells to MPT0B098 cytotoxicity, indicating that STAT3 is a key mediator of drug resistance in oral carcinogenesis. Moreover, the combination of MPT0B098 with the clinical drug cisplatin or 5-FU significantly augmented growth inhibition and apoptosis in OSCC cells. Taken together, our results provide a novel mechanism for the action of MPT0B098 in which the JAK2/STAT3 signaling pathway is suppressed through the modulation of SOCS3 protein level. The findings also provide a promising combinational therapy of MPT0B098 for OSCC.

No MeSH data available.


Related in: MedlinePlus

MPT0B098 induces the cell cycle arrest and apoptosis.(A) OEC-M1 cells were treated with 0.25 or 0.5 μM of MPT0B098 for 12 hrs. Cells were then evaluated for effects on cell cycle using PI staining and analyzed by flow cytometry. Percentages of cells in different phases were shown. The data are representative of three independent experiments. (B) OEC-M1 cells were treated with different concentrations of MPT0B098 for 12 hrs. Apoptosis was assessed by annexin V/PI staining and analyzed by flow cytometry. The data are represented as mean ± SE; ***, p<0.001 versus vehicle control. (C) OEC-M1 cells were incubated with various concentrations of MPT0B098 for 12~24 hours and caspases-3 activity was assessed. The data are represented as mean ± SE; **, p<0.01; ***, p<0.001 versus vehicle control. (D) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. The proteolytic cleavage of caspase-3 and PARP were determined by Western blot analysis. GAPDH was used as protein loading control. (E) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. Effects on the expression of Pim-1, Mcl-1, Bcl-2 and survivin were determined by western blot analysis. GAPDH was used as protein loading control.
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pone.0158440.g002: MPT0B098 induces the cell cycle arrest and apoptosis.(A) OEC-M1 cells were treated with 0.25 or 0.5 μM of MPT0B098 for 12 hrs. Cells were then evaluated for effects on cell cycle using PI staining and analyzed by flow cytometry. Percentages of cells in different phases were shown. The data are representative of three independent experiments. (B) OEC-M1 cells were treated with different concentrations of MPT0B098 for 12 hrs. Apoptosis was assessed by annexin V/PI staining and analyzed by flow cytometry. The data are represented as mean ± SE; ***, p<0.001 versus vehicle control. (C) OEC-M1 cells were incubated with various concentrations of MPT0B098 for 12~24 hours and caspases-3 activity was assessed. The data are represented as mean ± SE; **, p<0.01; ***, p<0.001 versus vehicle control. (D) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. The proteolytic cleavage of caspase-3 and PARP were determined by Western blot analysis. GAPDH was used as protein loading control. (E) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. Effects on the expression of Pim-1, Mcl-1, Bcl-2 and survivin were determined by western blot analysis. GAPDH was used as protein loading control.

Mentions: To determine the effects of MPT0B098 on cell cycle and apoptosis, OEC-M1 cells were treated with different concentrations of MPT0B098 and the cell cycle progression and the alteration of apoptotic markers in the cells were analyzed. As shown in Fig 2A, MPT0B098 induced a dose-dependent G2/M arrest at 12 h following the drug exposure. At the drug concentrations of 0.25 and 0.5 μmol/L, the population of OEC-M1 cells in G2/M phase was 46.03% and 61.53%, respectively (Fig 2A). In addition, exposure of MPT0B098 for 12 h also caused a dose-dependent induction of cell apoptosis as determined by annexin V-FITC staining but not PI staining (Annexin V+/PI-) (Fig 2B). Notably, dose-dependent increases in caspase-3 activation and PARP cleavage were also observed in MPT0B098-treated cells (Fig 2C and 2D). Moreover, the effect of MPT0B098 on the levels of anti-apoptotic proteins in OSCC cells was also investigated. As shown in Fig 2E, MPT0B098 caused a dose-dependent down-regulation of Bcl-2, Pim-1, Survivin, and Mcl-1 proteins. These findings suggest that in OSCC cells the induction of apoptosis by MPT0B098 may be mediated by its inhibition on the expression of these anti-apoptotic proteins.


MPT0B098, a Microtubule Inhibitor, Suppresses JAK2/STAT3 Signaling Pathway through Modulation of SOCS3 Stability in Oral Squamous Cell Carcinoma.

Peng HY, Cheng YC, Hsu YM, Wu GH, Kuo CC, Liou JP, Chang JY, Jin SL, Shiah SG - PLoS ONE (2016)

MPT0B098 induces the cell cycle arrest and apoptosis.(A) OEC-M1 cells were treated with 0.25 or 0.5 μM of MPT0B098 for 12 hrs. Cells were then evaluated for effects on cell cycle using PI staining and analyzed by flow cytometry. Percentages of cells in different phases were shown. The data are representative of three independent experiments. (B) OEC-M1 cells were treated with different concentrations of MPT0B098 for 12 hrs. Apoptosis was assessed by annexin V/PI staining and analyzed by flow cytometry. The data are represented as mean ± SE; ***, p<0.001 versus vehicle control. (C) OEC-M1 cells were incubated with various concentrations of MPT0B098 for 12~24 hours and caspases-3 activity was assessed. The data are represented as mean ± SE; **, p<0.01; ***, p<0.001 versus vehicle control. (D) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. The proteolytic cleavage of caspase-3 and PARP were determined by Western blot analysis. GAPDH was used as protein loading control. (E) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. Effects on the expression of Pim-1, Mcl-1, Bcl-2 and survivin were determined by western blot analysis. GAPDH was used as protein loading control.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4930189&req=5

pone.0158440.g002: MPT0B098 induces the cell cycle arrest and apoptosis.(A) OEC-M1 cells were treated with 0.25 or 0.5 μM of MPT0B098 for 12 hrs. Cells were then evaluated for effects on cell cycle using PI staining and analyzed by flow cytometry. Percentages of cells in different phases were shown. The data are representative of three independent experiments. (B) OEC-M1 cells were treated with different concentrations of MPT0B098 for 12 hrs. Apoptosis was assessed by annexin V/PI staining and analyzed by flow cytometry. The data are represented as mean ± SE; ***, p<0.001 versus vehicle control. (C) OEC-M1 cells were incubated with various concentrations of MPT0B098 for 12~24 hours and caspases-3 activity was assessed. The data are represented as mean ± SE; **, p<0.01; ***, p<0.001 versus vehicle control. (D) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. The proteolytic cleavage of caspase-3 and PARP were determined by Western blot analysis. GAPDH was used as protein loading control. (E) OEC-M1 cells were treated with different concentrations of MPT0B098 for 24 hrs. Effects on the expression of Pim-1, Mcl-1, Bcl-2 and survivin were determined by western blot analysis. GAPDH was used as protein loading control.
Mentions: To determine the effects of MPT0B098 on cell cycle and apoptosis, OEC-M1 cells were treated with different concentrations of MPT0B098 and the cell cycle progression and the alteration of apoptotic markers in the cells were analyzed. As shown in Fig 2A, MPT0B098 induced a dose-dependent G2/M arrest at 12 h following the drug exposure. At the drug concentrations of 0.25 and 0.5 μmol/L, the population of OEC-M1 cells in G2/M phase was 46.03% and 61.53%, respectively (Fig 2A). In addition, exposure of MPT0B098 for 12 h also caused a dose-dependent induction of cell apoptosis as determined by annexin V-FITC staining but not PI staining (Annexin V+/PI-) (Fig 2B). Notably, dose-dependent increases in caspase-3 activation and PARP cleavage were also observed in MPT0B098-treated cells (Fig 2C and 2D). Moreover, the effect of MPT0B098 on the levels of anti-apoptotic proteins in OSCC cells was also investigated. As shown in Fig 2E, MPT0B098 caused a dose-dependent down-regulation of Bcl-2, Pim-1, Survivin, and Mcl-1 proteins. These findings suggest that in OSCC cells the induction of apoptosis by MPT0B098 may be mediated by its inhibition on the expression of these anti-apoptotic proteins.

Bottom Line: In the present study, we examined the effect of a novel small-molecule microtubule inhibitor, MPT0B098, on STAT3 signaling in oral squamous cell carcinoma (OSCC).The accumulation of SOCS3 protein enhanced its binding to JAK2 and TYK2 which facilitated the ubiquitination and degradation of JAK2 and TYK2, resulting in a loss of STAT3 activity.Moreover, the combination of MPT0B098 with the clinical drug cisplatin or 5-FU significantly augmented growth inhibition and apoptosis in OSCC cells.

View Article: PubMed Central - PubMed

Affiliation: National Institute of Cancer Research, National Health Research Institutes, Miaoli, Taiwan.

ABSTRACT
Microtubule inhibitors have been shown to inhibit Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signal transduction pathway in various cancer cells. However, little is known of the mechanism by which the microtubule inhibitors inhibit STAT3 activity. In the present study, we examined the effect of a novel small-molecule microtubule inhibitor, MPT0B098, on STAT3 signaling in oral squamous cell carcinoma (OSCC). Treatment of various OSCC cells with MPT0B098 induced growth inhibition, cell cycle arrest and apoptosis, as well as increased the protein level of SOCS3. The accumulation of SOCS3 protein enhanced its binding to JAK2 and TYK2 which facilitated the ubiquitination and degradation of JAK2 and TYK2, resulting in a loss of STAT3 activity. The inhibition of STAT3 activity led to sensitization of OSCC cells to MPT0B098 cytotoxicity, indicating that STAT3 is a key mediator of drug resistance in oral carcinogenesis. Moreover, the combination of MPT0B098 with the clinical drug cisplatin or 5-FU significantly augmented growth inhibition and apoptosis in OSCC cells. Taken together, our results provide a novel mechanism for the action of MPT0B098 in which the JAK2/STAT3 signaling pathway is suppressed through the modulation of SOCS3 protein level. The findings also provide a promising combinational therapy of MPT0B098 for OSCC.

No MeSH data available.


Related in: MedlinePlus